A new trend to determine biochemical parameters by quantitative FRET assays.

Liao JY, Song Y, Liu Y - Acta Pharmacol. Sin. (2015)

Bottom Line:
Historically, FRET assays have been used to quantitatively deduce molecular distances.In this review, we discuss the use of quantitative FRET assays for the determination of biochemical parameters, such as the protein interaction dissociation constant (K(d)), enzymatic velocity (k(cat)) and K(m).We also describe fluorescent microscopy-based quantitative FRET assays for protein interaction affinity determination in cells as well as fluorimeter-based quantitative FRET assays for protein interaction and enzymatic parameter determination in solution.

Affiliation: Department of Bioengineering, University of California at Riverside, 900 University Avenue, Riverside, CA 92521, USA.

ABSTRACTFörster resonance energy transfer (FRET) has been widely used in biological and biomedical research because it can determine molecule or particle interactions within a range of 1-10 nm. The sensitivity and efficiency of FRET strongly depend on the distance between the FRET donor and acceptor. Historically, FRET assays have been used to quantitatively deduce molecular distances. However, another major potential application of the FRET assay has not been fully exploited, that is, the use of FRET signals to quantitatively describe molecular interactive events. In this review, we discuss the use of quantitative FRET assays for the determination of biochemical parameters, such as the protein interaction dissociation constant (K(d)), enzymatic velocity (k(cat)) and K(m). We also describe fluorescent microscopy-based quantitative FRET assays for protein interaction affinity determination in cells as well as fluorimeter-based quantitative FRET assays for protein interaction and enzymatic parameter determination in solution.

Mentions:
Before being digested by SENP1c, the total fluorescent emission of CyPet-(pre-SUMO1)-YPet at 530 nm in response to excitation at 414 nm can be divided into three parts: the FRET-induced emission of the acceptor (Ida), the direct emission of the donor (Id530/414) and the direct emission of the acceptor (Ia530/414), as shown in Figure 4:

Mentions:
Before being digested by SENP1c, the total fluorescent emission of CyPet-(pre-SUMO1)-YPet at 530 nm in response to excitation at 414 nm can be divided into three parts: the FRET-induced emission of the acceptor (Ida), the direct emission of the donor (Id530/414) and the direct emission of the acceptor (Ia530/414), as shown in Figure 4:

Bottom Line:
Historically, FRET assays have been used to quantitatively deduce molecular distances.In this review, we discuss the use of quantitative FRET assays for the determination of biochemical parameters, such as the protein interaction dissociation constant (K(d)), enzymatic velocity (k(cat)) and K(m).We also describe fluorescent microscopy-based quantitative FRET assays for protein interaction affinity determination in cells as well as fluorimeter-based quantitative FRET assays for protein interaction and enzymatic parameter determination in solution.

Affiliation:
Department of Bioengineering, University of California at Riverside, 900 University Avenue, Riverside, CA 92521, USA.

ABSTRACTFörster resonance energy transfer (FRET) has been widely used in biological and biomedical research because it can determine molecule or particle interactions within a range of 1-10 nm. The sensitivity and efficiency of FRET strongly depend on the distance between the FRET donor and acceptor. Historically, FRET assays have been used to quantitatively deduce molecular distances. However, another major potential application of the FRET assay has not been fully exploited, that is, the use of FRET signals to quantitatively describe molecular interactive events. In this review, we discuss the use of quantitative FRET assays for the determination of biochemical parameters, such as the protein interaction dissociation constant (K(d)), enzymatic velocity (k(cat)) and K(m). We also describe fluorescent microscopy-based quantitative FRET assays for protein interaction affinity determination in cells as well as fluorimeter-based quantitative FRET assays for protein interaction and enzymatic parameter determination in solution.